The purpose of this application is to further develop the candidate's skills in molecular and developmental biology towards attaining independent status as an investigator in cardiovascular disease and development. The candidate is currently an Assistant Professor in Cardiothoracic Surgery with a basic foundation in cardiac molecular biology. The work will be performed with strong institutional support and an environment that includes a strong basic research program in cardiovascular biology. The research plan will be performed within the community of the cardiovascular research institute of UCSF, under the mentors hip of Professor Charles Ordahl, a leader and expert in molecular and developmental biology of muscle differentiation. The aims of this project are to study the combinatorial interactions of cardiac promoter motifs and nuclear transcription factors that govern the regulation of cardiac-specific gene expression during development. Since unifying mechanisms probably govern the coordinate expression of cardiac genes during differentiation it is our hypothesis that specification of the cardiac phenotype requires the activity of "master" regulatory transcription factors. The avian cardiac troponin T promoter will be used as a model gene. The proposed research in this application involves (1) mutational analysis of promoter sequences believed to be essential for cardiac-specific gene expression (2) DNA-protein binding studies, proteolytic mapping and cloning to help identify known and novel transcription factors that activate cardiac promoters, and (3) transfection of single or multiple cardiac transcription factors in vitro and in vivo to activate cardiac-specific gene promoters. The role of transcriptional enhancer factor-1, a multigene family that we have recently cloned and characterized, in cardiac gene regulation will be investigated using a combination of molecular and classical embryological techniques. The identification of cardiac determination factors may be exploited for the development of bioengineered cardiac myoblasts for cellular transplantation in heart failure due to acquired and congenital heart disease. An understanding of the transcriptional mechanisms that control cardiogenesis may also elucidate the molecular pathogenesis of various congenital heart defects and cardiomyopathies.